We are studying the relationship between the detailed parameters of protein synthesis and the growth rate of Escherichia coli. After energy source shift-down, the rate of polypeptide chain propagation is unaltered, but the translational yield declines because of a reduced rate of translational initiation, leading to the accumulation of mRNA-ribosome initiation complexes. The rate of mRNA inactivation decreases with decreasing translational initiation. We propose to examine the intracellular pools of metabolic intermediates after a shift-down to identify the physiological effector(s) of translational control. The detailed structure of the iniation complexes, including changes in ribosomal protein and initiation factors, will be examined in order to identify the regulated step in initiation. Translational parameters will be measured in cells growing at various steady-state rates to determine if translational initiation and mRNA inactivation are quantitatively regulated as a function of growth rate. We will also use the initiation complexes produced during shift-down to isolate the fragment of mRNA which is protected by the ribosomes and determine whether these fragments, from a highly heterogeneous mRNA population, have any common oligonucleotide sequences which might serve as ribosome binding sites on mRNA. Because the presence of a ribosome bound to the 5'-end of mRNA confers functional stability on that mRNA, we hypothesize that mRNA inactivation involves a primary nuclease attack at or near the 5'-end. We propose to attempt isolation of such an endonuclease from E. coli and to study its specificity of attack on isolated mRNA and mRNA bound to single ribosomes in initiation complexes.